The superfamily of growth factor receptor tyrosine kinases has received a great deal of attention because of the suspected involvement of some of its members in the development of human cancers. Despite this attention, the molecular mechanisms that underlie many of the key events which accompany the activation of these receptors, and their subsequent interactions with (exogenous) phospho-substrates, have not been delineated. The overall goal of the proposed studies is to take advantage of the recent development of fluorescence spectroscopic approaches for monitoring ligand (growth factor)-receptor and receptor-receptor interactions, together with the availability of reconstituted growth factor receptor systems, and insect cell expression systems for different receptor tyrosine kinase domains, to characterize receptor-receptor and receptor-substrate interactions, and the receptor conformational changes accompanying these events within well defined reconstituted systems. A primary focus of these studies will be the members of the sub-group receptor tyrosine kinases; specifically, the epidermal growth factor (EGF) receptor, the neu/erbB-2 tyrosine kinase, and erbB-3. In many cases, the insulin receptor, which is the proto-type for the sub-group 2 family, also will be examined for comparative purposes. The proposed studies are sub- divided into four specific aims. 1.) Characterization of receptor (homo)dimer formation and its role in the activation of the EGF receptor and related receptor tyrosine kinases. 2.) Examination of the role of inter-receptor (trans) phosphorylation in the activation of the EGF receptor and related receptor tyrosine kinases. 3.) Characterization of the growth factor and metal ion activator-induced conformational changes within the tyrosine kinase domains of growth factor receptors. 4.) Reconstitution of the interactions of the EGF receptor with GTP-binding proteins. A number of specific questions will be addressed. These include whether the formation of stable receptor homodimers is pre-requisite for the generation of receptor tyrosine kinase activity, do intermolecular (trans) phosphorylation events between EGF receptor molecules or between the EGF receptor and neu/erbB-2 or erbB-3 result in kinase activation, and what types of conformational changes within the EGF receptor tyrosine kinase domain accompany kinase activation and do similar conformational changes accompany the activation of neu/erbB-2 by the recently identified neu ligand (Heregulin)? Other questions include how EGF-induced conformational changes compare with those induced by the glycoprotein, ASGP-2, which is capable of a biphasic regulation of the EGF receptor tyrosine kinase activity, and how do different GTP-binding proteins influence the activation of the EGF receptor or the ability of the activated receptor to bind phospho-substrates? An understanding on a molecular level of the different protein-protein interactions and accompanying conformational changes that underlie receptor tyrosine kinase-signaling will be pre-requisite for the development of strategies which will correct the signaling malfunctions responsible for the malignant state.